We have isolated and characterized the coding sequence of a novel human gene, termed arg, which is the second member of the Abelson subfamily of nonreceptor tyrosine kinases. The coding sequence and activated forms of it have been expressed in various systems to characterize its biological properties. To characterize the arg protein product, the arg coding sequence was expressed in bacteria using an IPTG inducible system. The recombinant protein was detected in bacterial lysates by immunoblotting and exhibited a molecular mass of 145 kDa. Phosphoamino acid analysis of the protein following an in vitro autokinase reaction revealed tyrosine phosphorylation and established that the arg protein cellular arg gene product was generated by expressing a carboxy terminal segment of the protein in bacteria and using the recombinant protein as an immunogen. The arg gene product was identified in cultured human cells as a 145- kDa protein that exhibited autokinase activity. Analysis of arg expression in murine tissues revealed that arg, like c-abl, is widely expressed, further extending the similarities between the two genes. A chimeric gag-arg molecule, similar to the gag-abl protein of v- abl, has been assembled and placed in a eukaryotic expression vector. Analysis of NIH/3T3 cells transfected with this vector revealed that, unlike gag-abl, it does not transform these cells. Phosphotyrosine analysis of the gag-arg protein revealed that it is less phosphorylated than the gag-abl counterpart. Further studies are underway to determine how the structure of these two related molecules affects their markedly different abilities to transform NIH/3T3 cells. In addition, the ability of the gag-arg molecule to transform hematopoietic cells is being tested.